Comparison of Different Progestagen Assays for Measuring Progesterone Metabolites in Faeces of the Bitch

Abstract

Summary From 12 bitches of various breeds with fertile oestrus cycles faecal samples were collected daily from the onset of pro‐oestrus till 20 days after the start of vulval bleeding, then once per week till about 1 week before term. Immunoreactive progesterone metabolites were extracted from the samples using methanol and measured using immunoassays. In a first experiment four different assays were compared in regard to the amounts of immunoreactive substances measured: the enzyme immunoassay against 20‐oxo‐3‐hydroxypregnanes showed twice to four times higher values of immunoreactive material than another using an antibody against 6β‐hydroxyprogesterone. An enzyme immunoassay for pregnanediol measured only low concentrations of immunoreactive material. Also a radio immunoassay using an antibody against 11β‐hyroxyprogesterone detected only small amounts of reacting material. High performance liquid chromatography showed that in faeces of bitches the immunoreactive progesterone metabolites were present in unconjugated form, mainly as 3α/β hydroxylated progestagens with a 20‐oxo group. In the second experiment the samples were measured with the assay system using the 20‐oxo‐3‐hydroxypregnane antibody. A few days before mating the concentration of progesterone metabolites increased, reaching 5.77 μmol/kg faeces (median) at the day of mating. High levels (10.45 μmol/kg faeces) were measured till the end of the first month after mating. Thereafter, the concentrations decreased, reaching 2.68 μmol/kg (median) at the end of the second month.

Characterization of reproductive cycles and adrenal activity in the black‐footed ferret (Mustela nigripes) by fecal hormone analysis

“…HPLC analysis suggested that fecal progestins in the black-footed ferret consist of a single conjugated and several unconjugated progestins, with native progesterone accounting for <10% of the total immunoreactivity [Brown, 1997]. The unconjugated progestins probably are reduced pregnanes, as has been described for other carnivores [Möstl et al, 1993;Brown et al, 1994;Monfort et al, 1997;Möstl and Brunner, 1997]. The antibody used in this EIA has been shown by HPLC to cross-react with several progestins that have similar retention times to pregnanes and hydroxyprogesterones in feces of the red wolf (Canis rufus) [Walker, 1999].…”

Behavioural and faecal glucocorticoid metabolite responses of single caging in six cats over 30 days

“…Samples were extracted and analysed according to Möstl and Palme's (2008) protocol using an enzyme immunoassay for 9.3 11-oxoaetiocholanolone (Lab-code: 72-alt, Palme and Möstl 1997 ) previously validated for use in cats ( Palme and others 2001 ). In the current study, interassay and intra-assay coefficients of variation were 8.9 per cent (n=12) and 3.5 per cent (n=383), respectively.…”

Fecal steroid analysis for monitoring reproduction in the sun bear (Helarctos malayanus)

“…In other carnivore species, i.e. the domestic dog, luteal phases were monitored analyzing fecal progesterone metabolites [27,28], whereas the pregnanediol assay used in our study on sun bears was found to be less suitable [28]. Additionally, in the dog, androgen metabolites are excreted into the feces during the follicular phase [27,29].…”